Strain KOS Research Articles

Structural relationship of regioselectively-sulfonated botryosphaeran derivatives on activity against herpes simplex virus type 1

The bioactivities of sulfonated polysaccharides are frequently related to their substitution pattern. In this study, the regioselective sulfonation of an exocellular fungal (1→3)(1→6)-β-D-glucan (botryosphaeran) was performed by two different methods: mild sulfonation (MS) and via pivaloyl ester (PS), in order to study the influence of the sulfonation pattern on the antiviral activity of the respective derivatives. Two sulfonated derivatives with substitution degrees of 0.82 (MS) and 0.49 (PS) were obtained, with substitution patterns at positions C-6, and C-2/C-4 of the glucose units, respectively. All derivatives were chemically characterized and evaluated for antiviral activity against Herpes simplex virus type 1 (HSV-1) KOS strain, and dengue type 2 (DENV-2). The sample sulfonated at positions C-6 (MS) showed a remarkable antiviral effect on HSV-1 (IC50 of 5.38 μg mL1), while PS remained inactive. The investigation of the mode of action of sample MS pointed to the inhibition of HSV-1 adsorption to the host cells. Both samples were inactive towards the dengue virus strain. This study demonstrated that the presence of sulfate groups at the C-6 positions of botryosphaeran is the preferred substitution pattern that enables the antiviral activity towards HSV-1.

Herpes Simplex Virus 1 Glycoprotein B from a Hyperfusogenic Virus Mediates Enhanced Cell-Cell Fusion.

Herpes simplex virus 1 (HSV-1) causes significant morbidity and death in humans worldwide. Herpes simplex virus 1 has a complex fusion mechanism that is incompletely understood. The HSV-1 strain ANG has notable fusion and entry activities that distinguish it from wild type. HSV-1 ANG virions fused with the Vero cell surface at 4 °C and also entered cells more efficiently at 15 °C, relative to wild type HSV-1 strain KOS virions, consistent with a hyperfusogenic phenotype. Understanding the molecular basis for the unique entry and fusion activities of HSV-1 strain ANG will help decipher the HSV fusion reaction and entry process. Sequencing of HSV-1 ANG genes revealed multiple changes in gB, gC, gD, gH, and gL proteins relative to wild type HSV-1 strains. The ANG UL45 gene sequence, which codes for a non-essential envelope protein, was identical to wild type KOS. HSV-1 ANG gB, gD, and gH/gL were necessary and sufficient to mediate cell-cell fusion in a virus-free reporter assay. ANG gB, when expressed with wild type KOS gD and gH/gL, increased membrane fusion, suggesting that ANG gB has hyperfusogenic cell-cell fusion activity. Replacing the KOS gD, gH, or gL with the corresponding ANG alleles did not enhance cell-cell fusion. The novel mutations in the ANG fusion and entry glycoproteins provide a platform for dissecting the cascade of interactions that culminate in HSV fusion and entry.

Inactivated herpes simplex virus-1 vaccine formulated in aqueous and alcoholic extracts of propolis boosts cellular and IgG responses.

In this study, the adjuvant activity of aqueous and alcoholic extracts of propolis was examined on the inactivated herpes simplex virus-1 (HSV-1). BALB/C mice were administered with inactivated (HSV-1; the KOS strain) plus alcoholic and aqueous extracts, followed by assessment of the cellular and humoral immune responses. Alcoholic and aqueous extracts, as an adjuvant, revealed a significant increase in lymphocyte proliferation and cytotoxic T lymphocyte (CTL) responses versus the HSV-1 group. In addition, HSV-1 plus alcoholic extract showed a remarkable increase in IFN-γ cytokine and IFN-γ/IL-4 ratio. On the other hand, both alcoholic and aqueous extracts in the HSV-1 vaccine suppressed the IL-4 cytokine response as compared with the HSV-1 vaccine. In addition, HSV-1 plus alcoholic extract showed a significant increment in IgG1, IgG2a, and IgG2b isotypes as compared with the HSV-1 vaccine. Propolis extracts seem to modulate the immune response against inactivated HSV-1 model and can be used as a suitable vaccine adjuvant or a component of a complex adjuvant against infectious diseases.

Anti-herpes activity of the total alkaloid fraction from the branches of Fusaea longifolia (Annonaceae)

ABSTRACT The herpes simplex virus type 1 (HSV-1) and type 2 (HSV-2) occur worldwide. Infections caused by these viruses have great public health importance due to the growing resistance to the first-choice drug, acyclovir, especially in immunosuppressed patients. Alkaloids derived from species of Annonaceae have been reported as antiviral agents against HSV and others viruses. Within this context, we evaluated the antiviral activity of the total alkaloid fraction (TAF) extracted from the branches of Fusaea longifolia (Aubl.) Saff. (Annonaceae), a species native to the Amazon region, against the HSV-1 and HSV-2 viruses. The antiviral activity was evaluated through the plate reduction assay and the mode of action was investigated by a set of other assays. The TAF was active against the HSV-2 strain 333 and against the HSV-1 strains KOS and 29R (acyclovir resistant), with selectivity index values (SI = 50% cytotoxic concentration/50% effective concentration) of 5, 4 and 3, respectively. In the preliminary study of the anti-HSV-2 mode of action, TAF showed viral inhibitory effects if added up to 12 h post-infection, had virucidal activity and did not present viral inhibition in pre-treatment. Our results showed that the TAF exhibited anti-HSV activity. Regarding HSV-2, TAF acted after the viral infection and had virucidal activity. A mass spectrometry analysis revealed the presence of nine alkaloids in the TAF that had previously been reported for Annonaceae, including liriodenine, lysicamine and isoboldine, which have been described as potential anti-HSV-1 agents.

HSV-1 LAT Promoter Deletion Viruses Exhibit Strain-Specific and LAT-Dependent Epigenetic Regulation of Latent Viral Genomes in Human Neurons.

Herpes simplex virus 1 (HSV-1) establishes latency in neurons and expresses long noncoding RNAs termed the latency-associated transcripts (LATs). Two previous studies using HSV-1 recombinants containing deletions in the LAT promoter revealed opposing effects of the promoter deletion regarding the heterochromatinization of latent viral genomes in mice ganglia. Confounding variables in these studies include viral strains utilized (17syn+ versus KOS), anatomical infection site (footpad versus eye) and infectious virus dose (500 versus 1 × 105 PFU), and to date the basis for the differences between the two studies remains unresolved. We recently reported that 17syn+ and KOS display distinct differences in heterochromatin levels during latency in human neurons. This raised the possibility that the discrepancy seen between the two previous studies could be explained by strain-specific differences within the LAT region. Here, we examine two recombinants containing orthologous 202 bp LAT promoter deletions, 17ΔPst and KOSΔPst, in a human neuronal model of latency and reactivation (LUHMES). We found that LUHMES neurons recapitulate previous observations in mice where deletion of the LAT promoter results in an increase in H3K27me3 deposition on the viral genome compared to the parental strain 17syn+ but a decrease compared to the parental strain KOS. We also found distinct strain-specific differences in the production of viral transcripts and proteins during latency. These results indicate that the function and/or regulation of the LATs differs between HSV-1 strains and may shed light on some discrepancies found in the literature when examining the function of the LATs. IMPORTANCE Herpes simplex virus 1 (HSV-1) establishes a lifelong infection in neuronal cells. Periodically, the virus reactivates from this latent state and causes recurrent disease. Mechanisms that control entry into and maintenance of latency are not well understood, though epigenetic posttranslational modification of histones associated with the viral genome are known to play an important role. During latency, the latency-associated transcript (LAT) is known to impact epigenetic marks, but the ultimate effect has been a point of controversy. Here, we utilize a human neuronal cell line model of HSV latency and reactivation (LUHMES) to characterize latency for two HSV-1 wild-type strains and their respective LAT promoter deletion viruses. We find that the LAT acts in a strain-specific manner to influence levels of chromatin marks, viral transcription, and viral protein production. This work highlights the need to account for strain-specific differences when characterizing the LAT's function and the dynamics of reactivation.

In vivo and in vitro per se effect evaluation of Polycaprolactone and Eudragit® RS100-based nanoparticles

Biodegradable polymeric nanocapsules (NC) present incredible characteristics as drug nanocarriers that optimize drug targeting. However, However, a more detailed isolated effect of polymer-based nanoparticles as drug carriers is required. This work aimed to evaluate the per se effect of blank-NC (NC-B) with different surface characteristics both in vitro and in vivo toxicity. NC1-B (Polysorbate 80 coated poly(ɛ-caprolactone) NC), NC2-B (polyethylene glycol 6000 coated poly(ɛ-caprolactone) NC), NC3-B (chitosan-coated poly(ɛ-caprolactone) NC) and NC4-B (Eudragit® RS100 NC) were prepared by nanoprecipitation method. Formulations were characterized by particle size, zeta potential, and pH. The in vitro cytotoxicity tests against tumor cell lines were performed (HepG2 and MCF-7). Antiviral activity was evaluated by MTT in Vero cells infected with HSV-1 (KOS strain). In vivo evaluation was performed in apomorphine-induced stereotypy in Wistar rats and locomotor activity distance, head movements, and rearing behavior were measured. NC1-B, NC2-B, NC3-B, and NC4-B had a diameter under 350 nm. The pH and zeta potential of formulations varied according to their coating. For in vitro evaluation of antitumor activity and antiviral activity, one-way ANOVA showed no significant differences in cell viability. In vivo tests showed low neurological effects. In conclusion, different surface characteristics of NC-B did not demonstrate toxicity against the evaluated cell lines HepG2 and MCF-7, antiviral effect against HSV-1, and the neurological effects in a stereotyping model were low and may be attributed to the per se effect of NC-B.

Natural Polyphenol-Containing Gels against HSV-1 Infection: A Comparative Study.

Herpes simplex virus type 1 infection commonly affects many people, causing perioral sores, as well as severe complications including encephalitis in immunocompromised patients. The main pharmacological approach involves synthetic antiviral drugs, among which acyclovir is the golden standard, often leading to resistant virus strains under long-term use. An alternative approach based on antiviral plant-derived compounds, such as quercetin and mangiferin, demonstrated an antiviral potential. In the present study, semisolid forms for cutaneous application of quercetin and mangiferin were designed and evaluated to treat HSV-1 infection. Phosphatidylcholine- and poloxamer-based gels were produced and characterized. Gel physical–chemical aspects were evaluated by rheological measurements and X-ray diffraction, evidencing the different thermoresponsive behaviors and supramolecular organizations of semisolid forms. Quercetin and mangiferin diffusion kinetics were compared in vitro by a Franz cell system, demonstrating the different gel efficacies to restrain the polyphenol diffusion. The capability of gels to control polyphenol antioxidant potential and stability was evaluated, indicating a higher stability and antioxidant activity in the case of quercetin loaded in poloxamer-based gel. Furthermore, a plaque reduction assay, conducted to compare the virucidal effect of quercetin and mangiferin loaded in gels against the HSV-1 KOS strain, demonstrated the suitability of poloxamer-based gel to prolong the polyphenol activity.

Activity of 3,19-isopropylidinyl andrographolide against herpes simplex virus type 1 in an animal model.

BackgroundIn our previous study, the semi-synthetic analog of andrographolide, 3,19-isopropylideneandrographolide (IPAD), acts more effectively against herpes simplex virus (HSV) infection in cell culture than does acyclovir. IPAD inhibits cytopathic effect and production of HSV wild types and drug-resistant strains. Its effect is associated with the reduction of immediate-early regulatory protein (ICP27) and early proteins (ICP8 and UL42), indicating a mode of action different from that of acyclovir. Therefore, studies of the anti-HSV activity of IPAD in animal models are required before further application.Material & MethodPrednisolone-treated BALB/c mice were cutaneously infected with HSV-1 wild-type KOS strain. Experimental groups included control groups (untreated or treated only with the cream base) and treatment groups (with acyclovir or IPAD creams). Creams were applied four times daily for 10 days after infection to the relevant groups. The skin lesion score was assessed twice a day for 10 days. In addition, the effect of IPAD on HSV copy number and HSV late gene (gD) expression was investigated in skin lesion cells by quantitative real-time polymerase chain reaction.ResultIPAD cream was significantly effective in delaying the development of skin lesions and regression of the skin lesion score by day 5 (P < 0.01) compared with untreated controls. In addition, this IPAD cream significantly reduced HSV DNA copy number and gD gene expression (P < 0.01). No signs of irritation were observed at the application site.ConclusionTopical administration of IPAD cream reduced skin lesions in mice cutaneously infected with HSV-1 KOS.

Adenovirus-mediated RNA interference against herpes simplex virus infection in vitro.

Herpetic keratitis caused by the herpes simplex virus (HSV) is the most common form of ocular herpes that causes corneal blindness. Although treatments for herpes keratitis have improved in recent years. there is still considerable room for new treatments against viral infection that shows great promise. The aim of the study was to evaluate the effect of RNA interference on HSV Type 1 (HSV1) infection in vitro, first prophylactically then therapeutically. The highly conserved glycoproteins D (gD) and E (gE) were chosen as targets for this study. Different small interfering RNA (siRNA) duplexes that target gD and gE were designed and chemically synthesized. The recombinant adenovirus type 5 was developed and used as the vehicle with which we delivered the siRNA into the Vero cells infected with the HSV1 KOS strain. Evaluation of the efficacy of siRNA-mediated inhibition was performed either before virus inoculation (prophylactically) or after virus inoculation at the first appearance of lesions (therapeutically). The expression of messenger RNA encoding gD and gE was detected using a real-time polymerase chain reaction (qPCR). We analyzed HSV replication in Vero cells, cytotoxicity of HSV, and cell viability. When used prophylactically, the siRNA-targeting gD and gE created a more marked decrease in viral titer than when used therapeutically. The transfection of cells with recombinant adenovirus containing the siRNA expression cassette was associated with very low cytotoxicity. Adenovirus-mediated siRNA-targeting gD and gE genes effectively inhibit the replication of the HSV in Vero cells. In addition, these findings indicate that the prophylactic use of siRNA is far more effective at inhibiting HSV replication than the therapeutic use.

Essential role of M1 macrophages in blocking cytokine storm and pathology associated with murine HSV-1 infection.

Ocular HSV-1 infection is a major cause of eye disease and innate and adaptive immunity both play a role in protection and pathology associated with ocular infection. Previously we have shown that M1-type macrophages are the major and earliest infiltrates into the cornea of infected mice. We also showed that HSV-1 infectivity in the presence and absence of M2-macrophages was similar to wild-type (WT) control mice. However, it is not clear whether the absence of M1 macrophages plays a role in protection and disease in HSV-1 infected mice. To explore the role of M1 macrophages in HSV-1 infection, we used mice lacking M1 activation (M1-/- mice). Our results showed that macrophages from M1-/- mice were more susceptible to HSV-1 infection in vitro than were macrophages from WT mice. M1-/- mice were highly susceptible to ocular infection with virulent HSV-1 strain McKrae, while WT mice were refractory to infection. In addition, M1-/- mice had higher virus titers in the eyes than did WT mice. Adoptive transfer of M1 macrophages from WT mice to M1-/- mice reduced death and rescued virus replication in the eyes of infected mice. Infection of M1-/- mice with avirulent HSV-1 strain KOS also increased ocular virus replication and eye disease but did not affect latency-reactivation seen in WT control mice. Severity of virus replication and eye disease correlated with significantly higher inflammatory responses leading to a cytokine storm in the eyes of M1-/- infected mice that was not seen in WT mice. Thus, for the first time, our study illustrates the importance of M1 macrophages specifically in primary HSV-1 infection, eye disease, and survival but not in latency-reactivation.

Sulfonated and Carboxymethylated β-Glucan Derivatives with Inhibitory Activity against Herpes and Dengue Viruses.

The infection of mammalian cells by enveloped viruses is triggered by the interaction of viral envelope glycoproteins with the glycosaminoglycan, heparan sulfate. By mimicking this carbohydrate, some anionic polysaccharides can block this interaction and inhibit viral entry and infection. As heparan sulfate carries both carboxyl and sulfate groups, this work focused on the derivatization of a (1→3)(1→6)-β-D-glucan, botryosphaeran, with these negatively-charged groups in an attempt to improve its antiviral activity. Carboxyl and sulfonate groups were introduced by carboxymethylation and sulfonylation reactions, respectively. Three derivatives with the same degree of carboxymethylation (0.9) and different degrees of sulfonation (0.1; 0.2; 0.4) were obtained. All derivatives were chemically characterized and evaluated for their antiviral activity against herpes (HSV-1, strains KOS and AR) and dengue (DENV-2) viruses. Carboxymethylated botryosphaeran did not inhibit the viruses, while all sulfonated-carboxymethylated derivatives were able to inhibit HSV-1. DENV-2 was inhibited only by one of these derivatives with an intermediate degree of sulfonation (0.2), demonstrating that the dengue virus is more resistant to anionic β-D-glucans than the Herpes simplex virus. By comparison with a previous study on the antiviral activity of sulfonated botryosphaerans, we conclude that the presence of carboxymethyl groups might have a detrimental effect on antiviral activity.

Mangiferin-Loaded Smart Gels for HSV-1 Treatment.

Infections due to HSV-1 affect many people all over the world. To counteract this pathology, usually characterized by perioral sores or by less frequent serious symptoms including keratitis, synthetic antiviral drugs are employed, such as acyclovir, often resulting in resistant viral strains under long-term use. Many plant-derived compounds, such as mangiferin and quercetin, have demonstrated antiviral potentials. In this study, smart semisolid forms based on phosphatidylcholine and Pluronic were investigated as delivery systems to administer mangiferin on skin and mucosae affected by HSV-1 infection. Particularly, lecithin organogels, Pluronic gel, and Pluronic lecithin organogels were formulated and characterized. After the selection of gel compositions, physical aspects, such as rheological behavior, spreadability, leakage, and adhesion were evaluated, suggesting a scarce suitability of the lecithin organogel for topical administration. Mangiferin was efficiently included in all type of gels. An in vitro study based on the Franz cell enabled us to find evidence of the gel capability to control drug diffusion, especially in the case of Pluronic organogel, while an in vivo study conducted on human volunteers demonstrated the safeness of all of the gels after cutaneous administration. Furthermore, a plaque reduction assay demonstrated the virucidal effect of mangiferin loaded in a Pluronic gel and a Pluronic lecithin organogel against the HSV-1 KOS strain.

Efficient establishment of reactivatable latency by an acyclovir-resistant herpes simplex virus 1 thymidine kinase substitution mutant with reduced neuronal replication

Herpes simplex virus 1 causes recurrent diseases by reactivating from latency, which requires the viral thymidine kinase (TK) gene. An acyclovir-resistant mutation in TK, V204G, was previously repeatedly identified in a patient with recurrent herpetic keratitis. We found that compared with its parental strain KOS, a laboratory-derived V204G mutant virus was impaired in replication in cultured neurons despite little defect in non-neuronal cells. After corneal inoculation of mice, V204G exhibited defects in ocular replication that were modest over the first three days but severe afterward. Acute replication of V204G in trigeminal ganglia was significantly impaired. However, V204G established latency with viral loads as high as KOS and reactivated with high frequency albeit reduced kinetics. Acyclovir treatment that drastically decreased ocular and ganglionic replication of KOS had little effect on V204G. Thus, despite reduced neuronal replication due to impaired TK activity, this clinically relevant drug-resistant mutant can efficiently establish reactivatable latency.

Abstract PO089: Comparison of Two oHSV Vectors for the Treatment of Glioblastoma

Abstract Glioblastoma multiforme (GBM) is the most common human brain cancer. Despite a well-established standard of care, the 5-year mortality rate of GBM patients is 95%, highlighting the need for innovative therapeutic interventions. A variety of oncolytic viruses, including those derived from herpes simplex virus (oHSV), have been designed for GBM therapy, but early-phase clinical trials have reported few complete responses without evidence of durable anti-tumor immunity. Potential reasons for the lack of efficacy are limited vector potency (i.e., virulence) and the presence of a highly immunosuppressive tumor microenvironment (TME) comprised of few activated lymphocytes, large numbers of immunosuppressive myeloid cells (macrophages, myeloid derived suppressor cells [MDSCs], microglia), and an agglomerate of immunosuppressive cytokines (IL-10, VEGF, MIF, etc.). Herein we explore these obstacles by comparing the anti-tumor activity of two different oHSV designs and by arming oHSV with an immunomodulatory payload designed to encourage the development of anti-tumor immunity. We first compared the activities of an HSV-1 KOS strain derivative designated KG4:T124 or colloquially, 2A5B, and an F strain derivative designated rQNestin34.5v.1 (a similar oHSV, rQNestin34.5v.2, is currently in a phase I clinical trial for GBM). We evaluated viral entry, replication capacity, and cytotoxicity using both murine and human glioma cell lines in vitro. Viral entry was relatively similar between both vectors, but rQNestin34.5v.1 replicated more effectively and generally induced greater viral mediated cytotoxicity. In syngeneic mice, rQNestin34.5v.1 reduced orthotopic GL261N4 tumor burden compared to 2A5B, but neither virus impeded disease progression in the more pernicious CT2A model. Evaluation of either the GL261N4 or CT2A TME revealed that both viruses induced a high influx of tumor associated macrophages, a transient polymorphonuclear (PMN) cell population, but did not alter the frequency of monocytic MDSCs. As a strategy to enhance oHSV efficacy, we sought to differentiate the highly immunosuppressive MDSC population into pro-inflammatory macrophages using oHSV armed with IL-12, a cytokine that has been shown to provide this activity. We show that oHSV:IL-12 induced a significant increase in tumor-associated macrophages and increased the survival of animals bearing either tumor type. Although we observed a low percentage of complete responses with the presence of anti-tumor CTLs demonstrated by tumor rechallenge experiments, IL-12 alone was not sufficient to significantly improve overall long-term animal survival, suggesting that additional arming genes should be considered in combination with IL-12. Citation Format: Joseph W. Jackson, Bonnie L. Hall, Lisa Bailey, Christophe Quéva, E. Antonio Chiocca, Justus B. Cohen, Joseph C. Glorioso. Comparison of Two oHSV Vectors for the Treatment of Glioblastoma [abstract]. In: Abstracts: AACR Virtual Special Conference: Tumor Immunology and Immunotherapy; 2020 Oct 19-20. Philadelphia (PA): AACR; Cancer Immunol Res 2021;9(2 Suppl):Abstract nr PO089.

HSV-1 Hijacks the Host DNA Damage Response in Corneal Epithelial Cells through ICP4-Mediated Activation of ATM

PurposeHerpes simplex virus type I (HSV-1) infection of corneal epithelial cells activates ataxia telangiectasia mutated (ATM), an apical kinase in the host DNA damage response pathway, whose activity is necessary for the progression of lytic HSV-1 infection. The purpose of this study is to investigate the mechanism of ATM activation by HSV-1 in the corneal epithelium, as well as its functional significance.MethodsMechanistic studies were performed in cultured human corneal epithelial cell lines (hTCEpi, HCE), as well as in esophageal (EPC2) and oral (OKF6) cell lines. Transfection-based experiments were performed in HEK293 cells. HSV-1 infection was carried out using the wild-type KOS strain, various mutant strains (tsB7, d120, 7134, i13, n208), and bacterial artificial chromosomes (fHSVΔpac, pM24). Inhibitors of ATM (KU-55933), protein synthesis (cycloheximide), and viral DNA replication (phosphonoacetic acid) were used. Outcomes of infection were assayed using Western blotting, qRT-PCR, immunofluorescence, and comet assay.ResultsThis study demonstrates that HSV-1-mediated ATM activation in corneal epithelial cells relies on the viral immediate early gene product ICP4 and requires the presence of the viral genome in the host nucleus. We show that ATM activation is independent of viral genome replication, the ICP0 protein, and the presence of DNA lesions. Interestingly, ATM activity appears to be necessary at the onset of infection, but dispensable at the later stages.ConclusionsThis study expands our understanding of HSV-1 virus-host interactions in the corneal epithelium and identifies potential areas of future investigation and therapeutic intervention in herpes keratitis.

Biological activities of marine invertebrates extracts from the northeast brazilian coast.

This paper reports the in vitro antiproliferative effects, antiprotozoal, anti-herpes and antimicrobial activities of 32 organic extracts of 14 marine sponges and 14 corals collected in northeast Brazilian coast. The ethanolic extracts of the sponges Amphimedon compressa and Tedania ignis, and the acetone extract of Dysidea sp. showed relevant results concerning the antiproliferative effects against A549, HCT-8, and PC-3 cell lines by sulforhodamine B assay, but also low specificity. Concerning the antiprotozoal screening, the ethanolic extract of Amphimedon compressa and the acetone and ethanolic extracts of Dysidea sp. were the most active against Leishmania amazonensis and Trypanosoma cruzi expressing β-galactosidase in THP-1 cells. In the preliminary anti-HSV-1 (KOS strain) screening, the ethanolic extracts of the sponges Amphimedon compressa, Haliclona sp. and Chondrosia collectrix inhibited viral replication by more than 50%. The most promising anti-herpes results were observed for the ethanolic extract of Haliclona sp. showing high selective indices against HSV-1, KOS and 29R strains (SI> 50 and >79, respectively), and HSV-2, 333 strain (IS>108). The results of the antibacterial screening indicated that only the ethanolic extract of Amphimedon compressa exhibited a weak activity against Enterococcus faecalis, Pseudomonas aeruginosa and Escherichia coli by the disk diffusion method. In view of these results, the extracts of Amphimedon compressa, Tedania ignis and Dysidea sp. were selected for further studies aiming the isolation and identification of the bioactive compounds with antiproliferative and/or antiprotozoal activities. The relevant anti-herpes activity of the ethanolic extract of Haliclona sp. also deserves special attention, and will be further investigated.

Mangiferin: A promising natural xanthone from Mangifera indica for the control of acyclovir – resistant herpes simplex virus 1 infection

Mangiferin is found in many plant species as the mango tree (Mangifera indica) with ethnopharmacological applications and scientific evidence. The emergence of resistant herpes simplex virus (HSV) strains to Acyclovir (ACV) has encouraged the search for new drugs. We investigated the in vitro and in vivo activity of mangiferin obtained from M. indica against ACV-resistant HSV-1 (AR-29) and sensitive (KOS) strains. The in vitro activity was performed under varying treatment protocols. The substance showed a CC50 > 500 μg/mL and IC50 of 2.9 μg/mL and 3.5 μg/mL, respectively, for the AR-29 and KOS strains. The in vivo activity was performed in Balb/c mice treated with 0.7% topical mangiferin formulation. This formulation inhibited most effectively the AR-29 strain, attenuated the lesions, postponed their appearance or enhanced healing, in comparison to control group. We demonstrated the potentiality of mangiferin from M. indica to control HSV replication with emphasis to ACV-resistant infection.

Identification and Pathogenic Potential of Orofacial Herpetic Clinical Isolates in Northeast Mexico

<i>Background</i> Herpes simplex viruses 1 (human herpes virus types 1, HSV-1) often cause recurrent infections that affect the skin, mouth, lips, and eyes and eventually induce herpetic encephalitis. A high percentage of the population is infected with HSV-1 in which it produces a variety of these orofacial disease. In Mexico, there are no studies to determine the effects of viral virulence of clinical facial dermal isolates of active infections caused by the Herpes simplex virus 1. <i>Objective</i> of this work was to compare the herpetic activity of human clinical isolates from northeast Mexico against HSV-1 KOS as reference strain, which induces experimental murine model keratitis disease produced by infecting mouse corneas. <i>Methods and Materials</i> we compared several clinical isolate of HSV-1 obtained from 25 patients diagnosed with HSV-1 active, according to acyclovir (ACV) susceptibility, thymidine kinase (TK) polymerase chain reaction (PCR), and experimental Balb/c mice model as viral infections <i>in vivo</i> were evaluated. Results we found that several clinical isolates showed ACV resistance (48%) and pathogenic potential (PP) differences that caused ocular infection more or less than reference HSV-1 KOS strain. In <i>Conclusion</i>, some clinical isolate from northeast Mexico shown differences that caused ocular infection more or less than reference HSV-1 KOS strain.

Functional Domains of the Herpes Simplex Virus Type 1 Tegument Protein pUL37: The Amino Terminus is Dispensable for Virus Replication in Tissue Culture.

The herpes simplex virus type 1 (HSV-1) UL37 gene encodes for a multifunctional component of the virion tegument, which is necessary for secondary envelopment in the cytoplasm of infected cells, for motility of the viral particle, and for the first steps in the initiation of virus infection. This 120 kDa protein has several known viral interacting partners, including pUL36, gK/pUL20, pUS10, and VP26, and cellular interacting proteins which include TRAF6, RIG-I, and dystonin. These interactions are likely important for the functions of pUL37 at both early and late stages of infection. We employed a genetic approach to determine essential domains and amino acid residues of pUL37 and their associated functions in cellular localization and virion morphogenesis. Using marker-rescue/marker-transfer methods, we generated a library of GFP-tagged pUL37 mutations in the HSV-1 strain KOS genome. Through viral growth and ultra-structural analysis, we discovered that the C-terminus is essential for replication. The N-terminal 480 amino acids are dispensable for replication in cell culture, although serve some non-essential function as viral titers are reduced in the presence of this truncation. Furthermore, the C-terminal 133 amino acids are important in so much that their absence leads to a lethal phenotype. We further probed the carboxy terminal half of pUL37 by alanine scanning mutagenesis of conserved residues among alphaherpesviruses. Mutant viruses were screened for the inability to form plaques—or greatly reduced plaque size—on Vero cells, of which 22 mutations were chosen for additional analysis. Viruses discovered to have the greatest reduction in viral titers on Vero cells were examined by electron microscopy (EM) and by confocal light microscopy for pUL37–EGFP cellular localization. This genetic approach identified both essential and non-essential domains and residues of the HSV-1 UL37 gene product. The mutations identified in this study are recognized as significant candidates for further analysis of the pUL37 function and may unveil previously undiscovered roles and interactions of this essential tegument gene.

Antiherpetic Effect of Topical Formulations Containing Sulfated Polysaccharide from Adenanthera pavonina.

Adenanthera pavonina is a native tree of Africa and Asia, introduced in Brazil for reforestation and wood industry. Several pharmacological activities have described scientifically, including antiviral activity. This study evaluated the antiviral effect of sulfated polysaccharide of Adenanthera pavonina (SPAp) against acyclovir (ACV)-resistant (AR-29) and sensitive (KOS) herpes simplex virus strains. The 50% cytotoxic concentration (CC50) was determined by MTT method and the 50% inhibitory concentration (IC50) was evaluated by plaque reduction assay. The in vivo SPAp antiviral activity was performed in Balb/c mice infected by skin scarification and treated with topical 0.5% (w/w) SPAp formulations. SPAp showed a CC50 of 47.81μg/mL and the IC50 were 0.49μg/mL (SI = 97.5) and 0.54μg/mL (SI = 88.5) for the strains KOS and AR-29, respectively. Our results demonstrated that mice treated with SPAp presented a delay in the development and progression of skin lesions compared with the control group.